Genome change is the result of functional interactions between proteins that maintain genome structure and the chromosomal structures present. An understanding of the mechanisms underlying these functional interactions is fundamental to understanding evolution, population variation, and somatic individuality. In previous work, we found that the chromosomal features correlating with specific gene deletion vulnerabilities include the presence or absence of a homologous chromosome, genome ploidy, and centromere environment. The proposed aims will convert these correlations to molecular mechanisms amenable to study in the yeast model, with relevance to future work in humans and other organisms. Our data suggest that specific genes acting in DNA repair and chromatin modification are particularly important in the absence of a homolog. In AIM 1, we propose to define the homolog interaction pathway(s). This may be especially important in outbred populations like our own, where chromosome structural variation includes frequent insertion/deletion difference between homologs. These structural variants may serve as enhancers of rearrangement in mutant cells. In our preliminary data, mutants that exhibit strong instability phenotypes only in diploid (but not haploid) cells may identify genes with importance proportional to chromosome number. In AIM 2, we will address this hypothesis. This pathway may reveal functions that are overwhelmed in cancer cells of high chromosome number. This may contribute to ongoing instability observed in such cells. Finally, centromere vulnerabilities tolerated well in wild-type cells are revealed in mutants. Yeast centromeres are divergent in DNA sequence, within the kinetochore-protein-binding regions and in surrounding chromatin context. It is likely that different viable mutants, which decrease segregation fidelity, affect centromeres differentially. In AIM 3, we will determine whether this is observed. Centromere evolution requires coordinate change in DNA sequence and binding proteins. In AIMS, we will also define the tolerated range of functional divergence through investigation of natural centromere fidelity as well as the genomic response to stringent challenge. In humans, distinct chromosome nondisjunction signatures that reveal specific dysfunctional pathways could enhance diagnosis and treatment of disease.